Restriction of Wnt signaling in the dorsal otocyst determines semicircular canal formation in the mouse embryo

The mouse inner ear develops from a simple epithelial pouch, the otocyst, with the dorsal and ventral portions giving rise to the vestibule and cochlea, respectively. The otocyst undergoes a morphological change to generate flattened saclike structures, known as outpocketings, in the dorsal and lateral regions. The semicircular canals of the vestibule form from the periphery of the outpocketings, with the central region (the fusion plate) undergoing de-epithelialization and disappearing. However, little is known of the mechanism that orchestrates formation of the semicircular canals. We now show that the area of canonical Wnt signaling changes dynamically in the dorsal otocyst during its morphogenesis. The genes for several Wnt ligands were found to be expressed in the dorsal otocyst according to specific patterns, whereas those for secreted inhibitors of Wnt ligands were expressed exclusively in the ventral otocyst. With the use of whole-embryo culture in combination with potent modulators of canonical Wnt signaling, we found that forced persistence of such signaling resulted in impaired formation both of the lateral outpocketing and of the fusion plates of the dorsal outpocketing. Canonical Wnt signaling was found to suppress Netrinl expression and to preserve the integrity of the outpocketing epithelium. In addition, inhibition of canonical Wnt signaling reduced the size of the otocyst, likely through suppression of cell proliferation and promotion of apoptosis. Our stage-specific functional analysis suggests that strict regulation of canonical Wnt signaling in the dorsal otocyst orchestrates the process of semicircular canal formation. (C) 2011 Elsevier Inc. All rights reserved.

小鼠内耳由一个简单的上皮囊袋——耳囊发育而来，其背侧和腹侧部分分别发育为前庭和耳蜗。耳囊经历形态变化，在背侧和外侧区域形成扁平的囊状结构，称为外突。前庭的半规管由外突的周边形成，而中心区域（融合板）发生上皮去分化并消失。然而，对于协调半规管形成的机制知之甚少。我们现在表明，在耳囊背侧的形态发生过程中，经典Wnt信号区域动态变化。发现几种Wnt配体的基因按照特定模式在耳囊背侧表达，而Wnt配体的分泌型抑制剂的基因仅在前庭腹侧表达。通过将全胚胎培养与经典Wnt信号的强效调节剂相结合，我们发现这种信号的强制持续存在会导致外侧外突以及背侧外突融合板的形成受损。发现经典Wnt信号可抑制Netrin1的表达并保持外突上皮的完整性。此外，抑制经典Wnt信号会减小耳囊的大小，这可能是通过抑制细胞增殖和促进细胞凋亡实现的。我们的阶段特异性功能分析表明，对耳囊背侧经典Wnt信号的严格调控协调了半规管形成的过程。（C）2011爱思唯尔公司。保留所有权利。